Project summary The capacity of the endoplasmic reticulum (ER) to fold and process proteins is maintained through a collection of signaling pathways termed the Unfolded Protein Response (UPR). The UPR is essential for development and differentiation of secretory cells and modulates a variety of human diseases including diabetes, cancer, and Alzheimer?s disease. The overall goal of this proposal is to understand the effects of the UPR in mammalian cells, focusing on two poorly understood aspects of its function: the regulation of mRNA decay and glucose metabolism. Activation of the mammalian UPR leads to the highly specific degradation of membrane- associated mRNAs. While this pathway has been observed in mice and is well-understood at the molecular level, its cell biological function is not clear. This proposal aims to understand the effects of this mRNA degradation pathway by focusing on its main target, Blos1. Both activation of the UPR and depletion of Blos1 by RNA interference lead to the perinuclear accumulation of lysosomes and the stabilization of Epidermal Growth Factor Receptor (EFGR) at the plasma membrane. The proposed research will use cultured cells to test the mechanism and consequences of these phenotypes, with the underlying hypothesis that lysosome re-positioning increases plasma membrane protein recycling, allowing cells to maintain surface protein activities by when trafficking from the ER is disrupted. The second goal of this proposal is to understand the interplay between the UPR and the metabolism of glucose. During UPR activation, cells switch their metabolism to aerobic glycolysis. This metabolic signature is also a hallmark of cancer cells, suggesting a role for the UPR in cancer metabolism. Hes1, a transcriptional repressor involved in differentiation, quiescence, and cancer, is a novel UPR target that down-regulates a key enzyme in the TCA cycle. This proposal will test the mechanism of metabolic regulation by the UPR, both in cells subjected to chemicals disrupting ER function and in cells differentiating into mature, collagen-secreting osteoblasts. The proposed experiments will then test two hypotheses regarding why the UPR regulates metabolism in this way: (a) that the reduction in oxidative phosphorylation protects cells from over-production of reactive oxygen species when ER oxidative protein folding is increased, and (b) that the upregulation of glycolysis is necessary to provide intermediates for increased glycoprotein and membrane production.